The present invention concerns the use of carbon-labeled aromatic amino acids, particularly phenylalanine and tyrosine, in a breath test for determining liver function. The test is exceptionally well-suited as a screening test for a variety of liver disorders including chronic liver disease such as cirrhosis, necrosis and hepatic degeneration due to hepatitis and other illnesses. The test can also provide quantitative information on liver function to identify the degree and progression of hepatic dysfunction. This information may be useful for planning liver transplants and other treatments.
Current tests for identification and management of liver dysfunction, are, primarily, static blood tests. These tests include measurements of specific enzyme activities such as aminotransferases or gamma glutamyltransferases, measurements of serum albumin, bile acids and bile pigments in plasma and urine, cholesterol testing, and coagulation tests such as prothrombin levels. These tests do not measure liver function but rather only show abnormal values which result from liver cell destruction. These tests are carried out primarily by blood testing in laboratories, although some stat instruments and methods for testing certain of these materials have been developed, or are in the process of development.
The problem with these static tests is that they do not provide a dynamic model of liver function but rather an indication of the prior liver cell destruction. Accordingly, in order to determine whether there are hepatic problems which could lead to the destruction of liver cells at a later time, a dynamic rather than static test is necessary.
Although a few specific quantitative tests for measuring hepatic function have been developed to supplement the static blood screening tests, these have not been overly successful. These dynamic tests are primarily breath tests measuring drug metabolism instead of degradation of natural metabolites. Examples of this first group of tests utilize labeled aminopyrine, phenacetin, or methacetin. Another group of breath tests, the substrate metabolism tests, use labeled galactose or caffeine. There are also plasma tests using caffeine and galactose. A further group of tests are plasma-based (as opposed to breath) liver substrate clearance tests using dyes such as indocyamine green and bromosulfophthalein.
The aminopyrine breath test has been found useful for measuring liver function in patients with alcoholic cirrhosis or chronic acute hepatitis. This test has clinical utility because of its capacity to reflect residual functional microsomal mass and viable hepatic tissue. See, e.g., A. Baker et al., Sem. in Liver Dis. 3:318-329 (1983). However, this test is useful mainly in identifying patients in most severe stages of liver disease. The reason is that the aminopyrine test is a measure of late function of cholestatic liver disease, not a sensor or indicator of early stages of disease. In addition, this test has potential safety concerns and significant cross reactivities which limit its usage.
Phenacetin and methacitin are also microsomal substrates which have been used in breath tests to evaluation hepatic function. Both phenacetin and methacetin undergo deethylation, resulting in acetaldehyde formation, which is oxidized by the Krebs cycle to CO.sub.2. However, the phenacetin and methacetin breath tests are not sensitive enough to evaluate mild liver disease.
The caffeine and galactose elimination breath tests are also useful in determining liver function but have problems of their own. The caffeine test is a quantitative measurement of hepatic microsomal metabolism while the galactose elimination test measures hepatic cytosolic function. The galactose breath test is a better discriminator of chronic liver damage then it is of mild liver damage but it is subject to diet interferences. Similarly, the caffeine test is inaccurate in cases of those who smoke or who have had environmental exposure of hazardous chemicals. The false positive results from this test, coupled with the very low CO.sub.2 values from healthy individuals, lead to too many inaccuracies. The plasma tests utilizing caffeine and galactose also share these same problems.
The dyes which are used in certain plasma tests have caused fatal reactions and accordingly have to be used under very controlled conditions. In addition, these are difficult tests to run and correlate with liver function.
A few papers have shown that plasma levels of labeled tyrosine or phenylalanine can be correlated, to some degree, with liver function. For example, Hehir et al., "Abnormal phenylalanine hydroxylation and tyrosine oxidation in a patient with acute fulminant liver disease with correction by liver transplantation," Gastroenterology 89: 659-65663 (1985), shows that in cases of chronic liver disease sufficiently acute so as to require transplantation, the use of isotope labeled phenylalanine and tyrosine could differentiate liver function values from those with properly functioning livers. Similarly, in Shanbhogue et al., "Whole body leucine, phenylalanine, and tyrosine kinetics in end-stage liver disease before and after hepatic transplantation", Metabolism 36( 11 ): 1047-1053 (1987), a test was made using three different labeled amino acids (phenylalanine, tyrosine and leucine) in order to detect hepatic dysfunction. Again, the primary tests used were plasma levels of each of these amino acids but some breath tests were also carried out. The breath tests yielded somewhat inconclusive results while the plasma test results were somewhat better. However, the study used patients with acute hepatic problems awaiting, and then undergoing, liver transplants. There is no indication that these tests could be used to measure liver function in non-acute patients.
Further, the Shanbhogue et al. paper used deuterated phenylalanine, .sup.13 C-leucine and .sup.14 C-tyrosine. The three labels were used to try to track the end-stage liver disease, not in any predictive function. The study was carried out using infusion rather than single intravenous or oral dose and the values determined were quantitative rather than being compared against a standard as in the present test. As such, there was no indication of what any breath values meant nor were they correlated in any way with clinical states. Specifically, the phenylalanine values showed no correlation for patients with liver disease or dysfunction. This is in direct contrast to the present test. The breath test in Shanbhogue et al. was not used as a predicitive test but merely as a means to obtain measurements of amino acid flux. Shanbhogue was directed to determining what changes occurred in plasma levels of these amino acids, not breath levels. As such, this work in no way affects the novelty or inventiveness of the present application.
Accordingly, an object of the invention is to provide a method of determining impairment of liver function at early stage of progression.
Another object of the invention is to provide a quick, inexpensive method of determining liver function.
A further object of the invention is to provide a screening test for liver function which can be used to detect both early and late liver disorders.
These and other objects and features of the invention will be apparent from the following description.